Role of Ascl1 in neuroblastoma tumorigenicity

FEDERICO JULIÁN GARDE; MARÍA JIMENA MOSNA; LUCÍA PAULA PETRIZ OTAÑO; MARCELO GABRIEL STINSON; DANIELA DI BELLA; ABEL LUIS CARCAGNO

Mar del plata

Congreso; LXVII Reunión Anual de la Sociedad Argentina de Investigación Clínica; 2022

SAIC

Neuroblastoma (NB) is the most common solid extracranial pediatric cancer derived from the sympathoadrenal lineage. Ascl1 is a proneural transcription factor whose transient expression is essential for the development of this lineage. Ascl1 remains overexpressed in NB cells and its high expression is associated with poor clinical prognosis. The aim of this work was to evaluate the role of Ascl1 in the tumorigenicity of NB cells. We evaluated the phenotype of Ascl1 knockdown (KD) SK-N-SH NB cells relative to control and analyzed scRNA-seq data from patients’ tumors to understand NB tumor cell heterogeneity. By performing a Trypan Blue assay, we found that Ascl1 KD does not affect cell viability. Analysis of proliferation by clonogenic assay and immunohistochemistry against Ki67 showed that the KD of Ascl1 significantly reduces the proliferating capacity of NB cells. Moreover, Ascl1 KD cells exhibited changes in their morphology relative to control, showing a neuron-like shape. For scRNA-seq analysis we obtained transcriptomic data of human NBs from public repositories. We integrated the data into a single dataset, performed dimensional reduction and clustering, and analyzed the origin of the cells. We characterized the resulting clusters through analysis of functional enrichment and analyzed expression of Ascl1 and co-expressed factors using non-negative matrix factorization. We found that NB cells share a single origin: sympathoblasts of the adrenal medulla, that cellular heterogeneity can be explained by the phase of the cell cycle, and that Ascl1 is expressed across all cell types in association with neurogenesis-related genes. The obtained results show that: a) Ascl1 blocks terminal neuronal differentiation in NBs, supporting tumor progression, b) a therapy against Ascl1 could target several tumoral subpopulations and, together with the observed effects of the Ascl1 KD in vitro sets the basis for a therapeutic strategy based on the modulation of Ascl1.